Biomedical subjects

R Bartrons

Publications and source records attributed to R Bartrons.

At least 73 records · Page 4Linked to original sources

PubMed1993

Fructose-1,6-bisphosphate fails to ameliorate delayed neuronal death in the CA1 area after transient forebrain ischaemia in gerbils.

Fructose-1,6-bisphosphate has been shown to reduce ischaemic-induced brain damage in rabbits and gerbils. In view of these findings, we investigated the effects of fructose-1,6-bisphosphate on delayed neuronal death, following bilateral forebrain ischaemia, in the gerbil hippocampus at the fourth day of reperfusion. We subjected gerbils to bilateral forebrain ischaemia for 20 min. Fructose-1,6-bisphosphate was administered: intraperitoneally at a dose of 1 g/kg in saline in hr before the occlusion or at a dose of 1 g/kg 1 hr before the occlusion and every 24 hr for 3 days; or intraventricularly at a dose of 0.1 g/kg just after the carotid occlusion. No significant differences in the number of dying cells in the CA1 area were found between each group of treated animals when compared with controls. This study suggests that fructose-1,6-bisphosphate, administered according to these three different schedules, fails to ameliorate delayed neuronal death after 20 min of bilateral forebrain ischaemia in the CA1 area of the gerbil hippocampus.

Animals↗

PubMed1993

Assessment of purine metabolism in human renal transplantation.

Cortical levels of nucleotides and their degradation products from 42 transplanted human kidneys have been studied. Biopsies were performed during renal harvesting just before cooling, at the end of cold storage, and following reinstallment of renal blood circulation. ATP levels fell, and AMP and degradation products (inosine monophosphate [IMP], inosine, adenosine, and hypoxanthine) increased during cold storage and returned to near-normal values 30 min after recirculation. The major degradation product found was hypoxanthine, indicating very poor xanthine oxidase activity in human kidneys. The sum of adenine nucleotides (ATP+ADP+AMP) did not significantly decrease after cold storage, but adenylate energy charge (ATP+1/2ADP/ATP+ADP+AMP) was reduced to half, being recovered in implanted kidneys. The sum of adenine nucleotides was significantly reduced after implantation. The rate of acute tubular necrosis was higher in kidneys preserved for more than 30 hr. Kidneys with acute tubular necrosis had significantly lower levels of the total pool of adenine nucleotides at reperfusion, but there was no correlation between incidence of acute tubular necrosis and ATP or other metabolite levels in the kidneys before or during cold preservation. The success of human kidney transplantation does not seem to depend only on the pool of residual nucleotides at the end of cold storage but on other factors that determine the ability of the cell to recover a normal energy state after reperfusion.

Adenosine↗

PubMed1992

Gene expression of regulatory enzymes of glycolysis/gluconeogenesis in regenerating rat liver.

Levels of mRNA for glucokinase, L-pyruvate kinase, fructose-1,6-bisphosphatase and phosphoenolpyruvate carboxykinase were analysed during liver regeneration. Levels of mRNA for glycolytic enzymes (glucokinase and L-pyruvate kinase) decreased rapidly after partial hepatectomy. Glucokinase mRNA increased at 16-24 h, returning to normal values after this time. L-pyruvate kinase mRNA recovered control levels at 168 h. In contrast, phosphoenolpyruvate carboxykinase mRNA increased rapidly after liver resection and remained high during the regenerative process. However, the levels of fructose-1,6-bisphosphatase mRNA were not modified significantly. These results correlate with the reported increased rate of gluconeogenesis and changes in enzyme levels after partial hepatectomy. The effect of stress on the mRNA levels was also studied. All enzymes showed variations in their mRNA levels after the surgical stress. In general, the differences were more pronounced in regenerating liver than in sham-operated animals, being practically normalized at 24 h.

Animals↗

PubMed1992

Bovine brain 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase. Evidence for a neural-specific isozyme.

Bovine brain 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase was purified to homogeneity and characterized. This bifunctional enzyme is a homodimer with a subunit molecular weight of 120,000, which is twice that of all other known bifunctional enzyme isozymes. The kinase/bisphosphatase activity ratio was 3.0. The Km values for fructose 6-phosphate and ATP of the 6-phosphofructo-2-kinase were 27 and 55 microM, respectively. The Km for fructose 2,6-bisphosphate and the Ki for fructose 6-phosphate for the bisphosphatase were 70 and 20 microM, respectively. Physiologic concentrations of citrate had reciprocal effects on the enzyme's activities, i.e. inhibiting the kinase (Ki of 35 microM) and activating the bisphosphatase (Ka of 16 microM). Phosphorylation of the brain enzyme was catalyzed by the cyclic AMP-dependent protein kinase with a stoichiometry of 0.9 mol of phosphate/mol of subunit and at a rate similar to that seen with the liver isozyme. In contrast to the liver isozyme, the kinetic properties of the brain enzyme were unaffected by cyclic AMP-dependent protein kinase phosphorylation, and also was not a substrate for protein kinase C. The brain isozyme formed a labeled phosphoenzyme intermediate and cross-reacted with antibodies raised against the liver isozyme. However, the NH2-terminal amino acid sequence of a peptide generated by cyanogen bromide cleavage of the enzyme had no identity with any known bifunctional enzyme sequences. These results indicate that a novel isozyme, which is related to other 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase isozymes, is expressed specifically in neural tissues.

Amino Acid Sequence↗

PubMed1992

Transcriptional and posttranscriptional regulation of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase during liver regeneration.

The control of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase (PFK-2/FBPase-2; EC 2.7.1.105/3.1.3.46) gene expression during liver regeneration was studied. The level of PFK-2/FBPase-2 mRNA decreased to about 5% of the control value 6 hr after partial hepatectomy. Thereafter the mRNA increased to a maximum at 48 hr and returned to normal levels by 96 hr. In sham-operated animals, only a small increase was observed during the first 4 hr. The mRNA was recognized by a 299-base-pair liver-specific cDNA probe but not by a muscle-specific probe. The time course of mRNA modulation was well correlated with PFK-2/FBPase-2 activity and with the amount of bifunctional enzyme protein determined by immunoblotting with an antibody raised against the N-terminal decapeptide of liver PFK-2/FBPase-2. No alteration in the degradation rate of PFK-2/FBPase-2 mRNA was noted after partial hepatectomy. The modulation of PFK-2/FBPase-2 gene expression during liver regeneration involved changes in the transcription rate. The rate decreased by 50% at 6 hr after liver resection. The rate increased thereafter to a maximum at 72 hr and then returned to control values by 96 hr. The transcription rate of albumin did not change, whereas that of phosphoenolpyruvate carboxykinase increased 12-fold at 6 hr. These results show that PFK-2/FBPase-2 gene transcription is specifically regulated and that this regulation is in part responsible for the alterations in hepatic metabolism seen in regenerating liver.

Animals↗

PubMed1992

Fructose 2,6-bisphosphate in developing rat brain.

Fructose 2,6-bisphosphate (Fru-2,6-P2) levels and 6-phosphofructo-1-kinase and 6-phosphofructo-2-kinase activities have been studied in rat brain during development from embryonal to adult state. Fru-2,6-P2 increases slightly from day 16 of gestation, reaching a maximum 24 h after birth, remaining quite constant during postnatal development. In contrast with 6-phosphofructo-1-kinase, which increases progressively after the first week of age, 6-phosphofructo-2-kinase remains unaltered throughout the period studied. The role of Fru-2,6-P2 in controlling cerebral glycolysis is discussed.

Animals↗

PubMed1992

Effect of galactosamine on hepatic carbohydrate metabolism: protective role of fructose 1,6-bisphosphate.

Intraperitoneal administration of galactosamine (400 mg/kg body wt) to rats results in reversible liver cell injury that is related to a dose-dependent depletion of uridine phosphates by formation of UDP-sugar derivatives. This damage was monitored through changes in serum enzymatic activities that increased after the first 6 hr of drug administration. Glycemia and serum albumin remained stable during liver injury, whereas cholesterol and triglycerides decreased. To maintain plasma glucose concentration, the hepatic carbohydrate metabolism was greatly altered. Glycogen dropped during the first hours, remaining low for up to 48 hr. Fructose 2,6-bisphosphate and ATP levels decreased even faster than glycogen, with lactate following a similar diminution and being restored in parallel with both metabolites. The reduction in fructose 2,6-bisphosphate can be explained by changes in the substrates or modulators of the 6-phosphofructo-2-kinase/fructose 2,6-bisphosphatase, because neither the cyclic AMP levels nor the activity ratio of the enzyme were modified. Simultaneous administration of galactosamine and fructose 1,6-bisphosphate (2 gm/kg) prevented liver cell death, as monitored by serum enzyme activities. Furthermore, the bisphosphorylated metabolite had protective effects on the changes in liver calcium content and ATP and fructose 2,6-bisphosphate concentrations. In contrast, fructose, fructose-1-phosphate and fructose-6-phosphate had no significant protection. Fructose 1,6-bisphosphate might decrease galactosamine toxicity by increasing fructose 2,6-bisphosphate and ATP levels, the changes in both metabolites probably being related. The significance of these findings with respect to the mechanism of galactosamine-induced liver injury is also discussed.

Adenosine Triphosphate↗

PubMed1992

Oral administration of vanadate to diabetic rats restores liver 6-phosphofructo-2-kinase content and mRNA.

Vanadate and insulin were administered to diabetic (streptozotocin) rats to compare their effects on the activity and mRNA content of 6-phosphofructo-2-kinase and L-type pyruvate kinase in the liver. The activity of 6-phosphofructo-2-kinase in livers of diabetic rats was about 40% of that found in normal rats. A similar decrease was found for 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase content, measured by immunoprecipitation, and for mRNA, measured by hybridization of Northern blots. Administration of vanadate to the diabetic rats led to a progressive recovery of 6-phosphofructo-2-kinase activity, and 6-phosphofructo-2-kinase/fructose- 2,6-bisphosphatase content and mRNA. This recovery, which was complete after 15 days of oral treatment, was also obtained after 60 h of insulin administration. L-type pyruvate kinase activity and mRNA were also decreased by about 70% in livers of diabetic rats. Both parameters normalized after 15 days of vanadate treatment, whereas insulin administration (60 h) raised L-pyruvate kinase mRNA three-fold above control values. Oral treatment for 15 days with vanadate can thus mimic the effect of insulin on both pyruvate kinase and 6-phosphofructo-2-kinase/fructose-2,6- bisphosphatase in livers of diabetic rats.

Administration, Oral↗

PubMed1992

HPLC analysis of hexosamine phosphates in biological samples.

Galactosamine is quickly metabolized to galactosamine 1-phosphate in rats treated with this compound. An HPLC method to quantify hexosamine phosphates in biological samples is described, modified from the o-phthaldialdehyde amino acid analysis procedure. o-Phthaldialdehyde derivatives of hexosamines and hexosamine-phosphates can be eluted from a reverse-phase column at different retention times, with a total analysis time of 30 min and without overlapping with free amino acids at physiological concentrations. The standard curves are linear between 1 and 40 nmol. This simple method is more selective and sensitive than previous enzymatic analyses of hexosamine phosphorylation.

Animals↗

PubMed1991

Effect of TPA on fructose 2,6-bisphosphate levels and protein kinase C activity in B-chronic lymphocytic leukemia (B-CLL).

Normal B lymphocytes and peripheral mononuclear blood cells from B-chronic lymphocytic leukemia (B-CLL) patients were incubated in the presence of the tumor promoting phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA). In normal B lymphocytes and lymphocytes from five patients with B-CLL, TPA stimulation increased lymphocyte fructose 2,6-bisphosphate (fructose 2,6-P2) content and activity of 6-phosphofructo 2-kinase (PFK-2), which is the enzyme that catalyzes the synthesis of fructose 2,6-P2. This effect was evident after 6 h and maximal after 12-24 h of TPA exposure. In three patients, lymphocytes seemed to be refractory to TPA stimulation in the conditions described here. Lymphocyte stimulation by TPA was associated with the translocation of protein kinase C (PKC) from the soluble to the particulate membrane fraction, except in B-CLL lymphocytes refractory to the TPA effect. These results give further support to the existence within B-CLL of subsets of cells which are refractory to the stimulation by TPA and demonstrate that the tumor promoter TPA induces important metabolic changes in lymphocytes of some patients with B-CLL.

B-Lymphocytes↗

PubMed1991

6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase in rat brain.

The concentration of fructose 2,6-bisphosphate in the brain remained stable during starvation and early stages of ischaemia, but decreased in diabetes or after lengthened ischaemia. 6-Phosphofructo-1-kinase activity was also decreased in diabetic and ischaemic animals, whereas 6-phosphofructo-2-kinase was not modified. The concentration of the bisphosphorylated metabolite seems to be remarkably constant under a wide variety of experimental conditions, suggesting that it plays an essential role in the basal activation of 6-phosphofructo-1-kinase. Purified 6-phosphofructo-2-kinase also showed fructose-2,6-bisphosphatase activity with an activity ratio similar to that of the purified heart isoenzyme. The brain enzyme also has a net charge similar to that of the heart isoenzyme. Its activity is not modified by sn-glycerol 3-phosphate, and it is more sensitive to citrate than the liver or muscle isoenzyme. Moreover, the enzyme from brain, similarly to that from heart and muscle, is not modified by the cyclic AMP-dependent protein kinase or protein kinase C. A near-full-length cDNA probe from liver hybridized with RNA from brain and heart. In both cases, a major band of 6.8 kb of RNA and a minor one of 4 kb of RNA were detected. All these properties support the hypothesis that brain contains a different isoenzymic form from that of liver and muscle, and it is probably related to the heart isoform.

Animals↗

PubMed1991

Vanadate induction of L-type pyruvate kinase mRNA in adult rat hepatocytes in primary culture.

In primary culture of adult rat hepatocytes, vanadate in the presence of glucose stimulates the expression of the liver (L-type) pyruvate kinase gene. Glucose by itself was inactive, and vanadate, like insulin, was also inefficient in the absence of glucose. Similar results were obtained on glucokinase gene expression. An analogue of cAMP, 8-(4-chlorophenylthio)-cAMP, inhibited the production of L-type pyruvate kinase and glucokinase mRNAs in the presence of glucose plus vanadate.

Animals↗

PubMed1990

Fructose 2,6-bisphosphate and 6-phosphofructo-2-kinase during liver regeneration.

Glycogen and fructose 2,6-bisphosphate levels in rat liver decreased quickly after partial hepatectomy. After 7 days the glycogen level was normalized and fructose 2,6-bisphosphate concentration still remained low. The 'active' (non-phosphorylated) form of 6-phosphofructo-2-kinase varied in parallel with fructose 2,6-bisphosphate levels, whereas the 'total' activity of the enzyme decreased only after 24 h, similarly to glucokinase. The response of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase from hepatectomized rats (96 h) to sn-glycerol 3-phosphate and to cyclic AMP-dependent protein kinase was different from that of the enzyme from control animals and similar to that of the foetal isoenzyme.

Animals↗

PubMed1990

Vanadate inhibits liver fructose-2,6-bisphosphatase.

Vanadate was found to be a reversible non-competitive inhibitor of chicken liver fructose-2,6-bisphosphatase. The inhibition was best observed in the presence of glycerol 2- or 3-phosphate and half-maximal effect was obtained with about 0.15 mM vanadate. Vanadate decreased the extent of phosphorylation of the enzyme (E-P) by fructose 2,6-[2-32P]bisphosphate. This did not result from an increased rate of E-P breakdown, as is the case with phosphoglycerate mutase, an enzyme which shares structural and functional similarity to fructose-2,6-bisphosphate. The data were consistent with the formation of a dead-end transition state analogue of phosphate in the active site. Inhibition of fructose-2,6-bisphosphatase by vanadate offers a likely explanation for the increase in fructose 2,6-bisphosphate concentration brought about by vanadate in isolated rat hepatocytes.

Animals↗

PubMed1990

The impairment of respiration by glycolysis in the Lewis lung carcinoma.

Isolated vegetative tumour cells from mice bearing the Lewis lung carcinoma showed low rates of basal respiration with both low oxygen uptake rates and cytochrome-c oxidase activity. The cells were affected by a marked Crabtree effect and a high rate of lactate production in the presence of 10 mM glucose. The glycolytic capacity of the tumour was also assessed through the measurement of the maximum activities for hexokinase, phosphofructokinase, pyruvate kinase and lactate dehydrogenase. These activities were similar to the ones found in other fast-growing, undifferentiated tumours. The concentration of fructose-2,6-bisphosphate in the tumour was 2,3 nmoles/g fresh tissue wt., a value which is of the same order of magnitude as that found in other types of highly glycolytic cells. It is concluded that the Lewis lung carcinoma follows the same pattern as other undifferentiated tumours with a high capacity for both glucose and amino acid utilization.

Animals↗

PubMed1990

Oral administration of vanadate to streptozotocin-diabetic rats restores the glucose-induced activation of liver glycogen synthase.

Isolated hepatocytes from streptozotocin-diabetic rats failed to respond to a glucose load with an activation of glycogen synthase. This lesion was associated with severely decreased activities of glycogen-synthase phosphatase and of glucokinase. All these defects were abolished after consumption for 13-18 days of drinking water containing Na3VO4 (0.7 mg/ml), and they were partially restored after 3.5 days, when the blood glucose concentration was already normalized. In all conditions the maximal extent of activation of glycogen synthase in cells closely parallelled the activity of glycogen-synthase phosphatase.

Animals↗

PubMed1990

Effects of vanadate on 6-phosphofructo 2-kinase activity and fructose 2,6-bisphosphate levels in cultured rat hepatocytes.

The presence of vanadate in primary cultures of rat hepatocytes produced a significant increase in the concentration of fructose 2,6-bisphosphate and in the activity of 6-phosphofructo 2-kinase. Compared with insulin, vanadate had a more potent action on the metabolite increase, but a similar effect on the 6-phosphofructo 2-kinase activity. Both the insulin- and the vanadate-dependent enhancements of 6-phosphofructo 2-kinase were inhibited by cycloheximide which specifically blocks protein synthesis on the translational level, suggesting that the increase of the enzyme activity was due to induction rather than to a change in the catalytic activity.

Animals↗

PubMed1990

Isolation and characterization of phosphofructo 2-kinase from chicken erythrocytes.

1. Phosphofructo 2-kinase from chicken erythrocytes copurifies with fructose 2,6-bisphosphatase activity, suggesting that the enzyme is bifunctional. 2. Similarly to phosphofructo 2-kinase from other tissues it is activated by inorganic phosphate, and inhibited by phosphoenol pyruvate, sn-glycerol 3-phosphate and citrate. However, it has some characteristics different than those of chicken liver phosphofructo 2-kinase, indicating that it is a distinct isozyme. 3. The phosphofructo 2-kinase/fructose 2,6-bisphosphatase activity ratio of the erythrocyte enzyme is one order of magnitude higher than that of the enzyme from liver. In contrast with the chicken liver enzyme, phosphofructo 2-kinase from chicken erythrocytes is activated by dithiothreitol and its activity increases with pH. 4. Chicken erythrocyte phosphofructo 2-kinase activity is neither modified by cyclic AMP-dependent protein kinase or casein kinase I and II. In contrast, it is partially inhibited by protein kinase C.

Animals↗